Filamentous morphology engineering of bacteria by iron metabolism modulation through MagR expression.

The morphology is the consequence of evolution and adaptation. Escherichia coli is rod-shaped bacillus with regular dimension of about 1.5 µm long and 0.5 µm wide. Many shape-related genes have been identified and used in morphology engineering of this bacteria. However, little is known about if specific metabolism and metal irons could modulate bacteria morphology. Here in this study, we discovered filamentous shape change of E. coli cells overexpressing pigeon MagR, a putative magnetoreceptor and extremely conserved iron-sulfur protein. Comparative transcriptomic analysis strongly suggested that the iron metabolism change and iron accumulation due to the overproduction of MagR was the key to the morphological change. This model was further validated, and filamentous morphological change was also achieved by supplement E. coli cells with iron in culture medium or by increase the iron uptake genes such as entB and fepA. Our study extended our understanding of morphology regulation of bacteria, and may also serves as a prototype of morphology engineering by modulating the iron metabolism.

Protective effect of Amauroderma rugosum ethanol extract and its primary bioactive compound, ergosterol, against acute gastric ulcers based on LXR-mediated gastric mucus secretions.

BACKGROUND: Amauroderma rugosum (Blume & T. Nees) Torrend (Ganodermataceae) is an edible mushroom with a wide range of medicinal values. Our previous publication demonstrated the therapeutic effects of the water extract of A. rugosum (WEA) against gastric ulcers. However, the protective effects of the ethanol extract of A. rugosum (EEA) on gastric mucosa and its major active constituents have not yet been elucidated. PURPOSE: This study aims to evaluate the gastroprotective effects and underlying mechanisms of EEA and its fat-soluble constituent, ergosterol, in acute gastric ulcers. STUDY DESIGN AND METHOD: SD rats were pre-treated with EEA (50, 100, and 200 mg/kg) or ergosterol (5, 10, and 20 mg/kg), and acute gastric ulcer models were constructed using ethanol, gastric mucus secretion inhibitor (indomethacin) or pyloric-ligation. The gastric ulcer area, histological structure alterations (H&E staining), and mucus secretion (AB-PAS staining) were recorded. Additionally, Q-PCR, western blotting, immunohistochemistry, ELISA, molecular docking, molecular dynamics simulations, MM-GBSA analysis, and surface plasmon resonance assay (SPR) were used to investigate the underlying mechanisms of the gastroprotective effect. RESULT: Compared with WEA, which primarily exerts its anti-ulcer effects by inhibiting inflammation, EEA containing fat-soluble molecules showed more potent gastroprotective effect through the promotion of gastric mucus secretion, as the anti-ulcer activity was partly blocked by indomethacin. Meanwhile, EEA exhibited anti-inflammatory effects by suppressing the production of IL-6, IL-1ß, TNF-α, and NO, thereby inhibiting the MAPK pathway. Significantly, ergosterol (20 mg/kg), the bioactive water-insoluble compound in EEA, exhibited a gastroprotective effect comparable to that of lansoprazole (30 mg/kg). The promotion of gastric mucus secretion contributed to the effects of ergosterol, as indomethacin can completely block it. The upregulations of COX1-PGE2 and C-fos, an activator protein 1 (AP-1) transcription factor, were observed after the ergosterol treatment. Ergosterol acted as an LXRß agonist via van der Waals binding and stabilizing the LXRß protein without compromising its flexibility, thereby inducing the upregulation of AP-1 and COX-1. CONCLUSION: EEA and its primary bioactive compound, ergosterol, exert anti-ulcer effects by promoting gastric mucus secretion through the LXRß/C-fos/COX-1/PGE2 pathway.

Sijunzi Decoction Targets IL1B and TNF to Reduce Neutrophil Extracellular Traps (NETs) in Ulcerative Colitis: Evidence from Silicon Prediction and Experiment Validation.

Purpose: This study was conducted to explore the mechanism of Sijunzi Decoction (SJZ) in the treatment of ulcerative colitis (UC). Methods: The study aimed to investigate the active components and targets of SJZ in the treatment of UC by screening databases such as TCMSP, GeneCards, OMIM, Distinct, TTD, and Drugbank. An online Venn tool, Cytoscape 3.7.2, and Autodock Tools were used to analyze the components and targets. The study also used a mouse model of UC to further investigate the effects of SJZ. HE staining, immunofluorescence, ELISA, qPCR, and Western blot were used to detect various indices. Results: Eighty-three active components and 112 action targets were identified from SJZ, including 67 targets for treating UC-related NETs. The five core targets identified were AKT1, JUN, IL1B, PTGS2, and TNF, and molecular docking studies indicated that the five targets were well-docked with ginsenoside Rh2, isoflavones, and formononetin. Animal experiments demonstrated that SJZ could alleviate various parameters such as weight, colon length, spleen index, disease activity index, and intestinal pathology of the UC mice. Immunofluorescence and Western blot showed that SJZ could reduce the expression of IL1B and TNF in intestinal neutrophils while increasing the expression of Occludin. Cellular immunofluorescence suggests that SJZ can reduce the expression of TNF and IL1B in NETs. The qPCR results also suggested that SJZ could inhibit TNF signal. Furthermore, ELISA results suggested that SJZ could inhibit the expression of pro-inflammatory cytokines (TNF-α, IL-1ß, IL-6) while promoting the expression of anti-inflammatory cytokines (IL-10, IL-37, TGF-ß). Conclusion: SJZ treats UC by reducing the content of intestinal NETs, with primary targets on the NETs being IL1B and TNFand suppress TNF signal. The practical components of SJZ may be ginsenoside Rh2, isoflavones, and formononetin.

Liuwei Dihuang Prevents Human Umbilical Vein Endothelial Cells Senescence via the JPX-STING-IRF3 Pathway.

BACKGROUND: Cellular senescence plays a crucial role in age-related diseases. Endothelial senescence is closely associated with age-related vascular disorders. This study aimed to reveal the role of traditional Chinese medicine Liuwei Dihuang (LWDH) in anti-endothelial cell senescence. METHODS: Human umbilical vein endothelial cells (HUVECs) were exposed to LPS treatment to induce senescence. Senescence-associated ß-galactosidase (SA-ß-gal) positive staining, p53 and p16 expression, BrdU staining, and relative telomere length (RTL) experiments were conducted to estimate LPS-induced cellular senescence of HUVECs. Real-time qPCR analysis was performed to identify differentially expressed lncRNAs in LPS-induced senescent HUVECs before and after treatment with LWDH. Bioinformatics analysis and ChIP assay were conducted to explore the mechanism of JPX in the anti-endothelial cell aging effect of LWDH. RESULTS: We first discovered that lncRNA JPX and STING-IRF3 pathways are involved in the process of anti-endothelial senescence of LWDH. Mechanistically, LWDH could reverse abnormally elevated JPX induced by LPS and inhibit the activation of STING, as well as the interaction between JPX and STING. CONCLUSION: Overall, our study explores the potential therapeutic value of LWDH and provides key insights into the potential avenues for preventing and treating HUVECs senescence.

A Rationally Designed Building Block of the Putative Magnetoreceptor MagR.

The ability of animals to perceive guidance cues from Earth's magnetic field for orientation and navigation has been supported by a wealth of behavioral experiments, yet the nature of this sensory modality remains fascinatingly unresolved and wide open for discovery. MagR has been proposed as a putative magnetoreceptor based on its intrinsic magnetism and its complexation with a previously suggested key protein in magnetosensing, cryptochrome, to form a rod-like polymer structure. Here, we report a rationally designed single-chain tetramer of MagR (SctMagR), serving as the building block of the hierarchical assembly of MagR polymer. The magnetic trapping experiment and direct magnetic measurement of SctMagR demonstrated the possibility of magnetization of nonmagnetic cells via overexpressing a single protein, which has great potential in various applications. SctMagR, as reported in this study, serves as a prototype of designed magnetic biomaterials inspired by animal magnetoreception. The features of SctMagR provide insights into the unresolved origin of the intrinsic magnetic moment, which is of considerable interest in both biology and physics. © 2022 Bioelectromagnetics Society.

The traditional herbal formulation, Jianpiyifei II, reduces pulmonary inflammation induced by influenza A virus and cigarette smoke in mice.

Chronic obstructive pulmonary disease (COPD) is a worldwide chronic inflammatory lung disease, and influenza A virus (IAV) infection is a common cause of acute exacerbations of COPD (AECOPD). Therefore, targeting viral infections represents a promising strategy to prevent the occurrence and development of inflammatory flare ups in AECOPD. Jianpiyifei II (JPYFII) is a traditional herbal medicine used in China to treat patients with COPD, and its clinical indications are not well understood. However, investigation of the anti-inflammatory effects and underlying mechanism using an animal model of smoking have been reported in a previous study by our group. In addition, some included herbs, such as Radix astragali and Radix aupleuri, were reported to exhibit antiviral effects. Therefore, the aim of the present study was to investigate whether JPYFII formulation relieved acute inflammation by clearing the IAV in a mouse model that was exposed to cigarette smoke experimentally. JPYFII formulation treatment during smoke exposure and IAV infection significantly reduced the number of cells observed in bronchoalveolar lavage fluid (BALF), expression of proinflammatory cytokines, chemokines, superoxide production, and viral load in IAV-infected and smoke-exposed mice. However, JPYFII formulation treatment during smoke exposure alone did not reduce the number of cells in BALF or the expression of Il-6, Tnf-a, and Il-1ß. The results demonstrated that JPYFII formulation exerted an antiviral effect and reduced the exacerbation of lung inflammation in cigarette smoke (CS)-exposed mice infected with IAV. Our results suggested that JPYFII formulation could potentially be used to treat patients with AECOPD associated with IAV infection.

A Systematic Review of the Mechanisms Underlying Treatment of Gastric Precancerous Lesions by Traditional Chinese Medicine.

Gastric precancerous lesions (GPLs) are an essential precursor in the occurrence and development of gastric cancer, known to be one of the most common and lethal cancers worldwide. Traditional Chinese medicine (TCM) has a positive prospect for the prevention and therapy of GPL owing to several advantages including a definite curative effect, fewer side effects compared to other treatments, multiple components, and holistic regulation. Despite these characteristic advantages, the mechanisms of TCM in treating GPL have not been fully elucidated. In this review, we summarize the current knowledge with respect to herbal formulations and the therapeutic mechanisms of TCM active ingredients for GPL. This paper elaborates on the mechanisms of TCM underlying the prevention and treatment of GPL, specifically those that are linked to anti-H. pylori, anti-inflammation, antiproliferation, proapoptotic, antioxidation, antiglycolytic, and antiangiogenesis effects.

The Neuroprotective Effects of Danggui-Shaoyao San on Vascular Cognitive Impairment: Involvement of the Role of the Low-Density Lipoprotein Receptor-Related Protein.

Effective drugs for treating dementia are still rare. Danggui-Shaoyao San (DSS), a traditional Chinese medicine, has been widely used in oriental countries for the treatment of various gynecological diseases. Many studies reported that DSS could ameliorate cognitive impairment. In this study, we aimed to investigate the underlying mechanism of DSS on vascular cognitive impairment (VCI) rats. Chronic cerebral hypoperfusion (CCH) is one of the main causes of VCI. CCH resulted in a chain of pathological process, including neuroinflammation, neuronal apoptosis, and oxidative stress. The most widely used animal model of VCI is permanent bilateral common carotid artery occlusion in rats. In this research, we determined whether DSS attenuated cognitive impairment by targeting I kappa B kinase (IKK)/nuclear factor of kappa B (NF-κB) signal pathway in VCI rats. Morris water maze and fear conditioning tests results indicated that DSS [7.2 g/(kg·d)] could improve learning and memory ability in VCI rats. We also found DSS significantly elevated the levels of low-density lipoprotein receptor-related protein 1 (LRP1) in the brain of VCI rats and this might indirectly target the IKK/NF-κB signal pathway to exert inhibitory effect on neuroinflammation, neuronal apoptosis, and oxidative stress in VCI rats. The present researches indicated that DSS might attenuate cognitive impairment by targeting IKK/NF-κB signal pathway in VCI rats and DSS might be a promising agent on VCI.

Cordyceps polysaccharide ameliorates airway inflammation in an ovalbumin-induced mouse model of asthma via TGF-ß1/Smad signaling pathway.

Allergic asthma is a chronic inflammatory disease characterized by airflow obstruction, airway hyperresponsiveness (AHR), airway inflammation, and mucus overproduction. Cordyceps polysaccharide (CPS) is one of the main bioactive compounds of Cordyceps militarisis, a traditional Chinese medicine. In this study, we established a mouse model of asthma using ovalbumin (OVA) challenge and evaluated the potential regulatory effect of CPS (25, 50, and 100 mg/kg) on asthmatic mice. These results showed that the asthmatic mice treated with CPS suppressed the secretion of eotaxin, IL-4, IL-5, IL-13, and IFN-γ in the blood and bronchoalveolar lavage fluid (BALF), and decreased serum IgE levels compared to the vehicle-treated mice. CPS also alleviated inflammatory cell infiltration, goblet cell hyperplasia, and the increases of inflammatory cells in the mouse model of asthma. In addition, OVA-induced AHR was inhibited by CPS treatment. Further analyses of protein expression revealed that CPS inhibited the activation of transforming growth factor ß1 (TGF-ß1)/Smad pathway in mice with asthma. These findings indicated that CPS might serve as a potential therapeutic agent for the management of allergic asthma.

Protective effects of Weipixiao decoction against MNNG-induced gastric precancerous lesions in rats.

Gastric cancer is recognized as one of the most common cancer. In-depth research of gastric precancerous lesions (GPL) plays an important role in preventing the occurrence of gastric cancer. Meanwhile, traditional treatment provides a novel sight in the prevention of occurrence and development of gastric cancer. The current study was designed to assess the effects of therapy with Weipixiao (WPX) decoction on N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-induced GPL rats and the underlying molecular mechanisms. After 10-weeks treatment, all rats were sacrificed. Histopathological changes of gastric tissue were assessed via hematoxylin-eosin (HE) and High-iron diamine-Alcian blue-Periodic acid-Schiff (HID-AB-PAS) staining. To be fully evidenced, RT-qPCR, Western blot and immunohistochemistry were used to detect the expressions of LDHA, CD147, HIF-1α, MCT4, PI3K, AKT, mTOR and miRNA-34a, which were crucial factors for evaluating GPL in the aspect of glycolysis pathogenesis. According to the results of HE and HID-AB-PAS staining, it could be confirmed that MNNG-induced GPL rats were obviously reversed by WPX decoction. Additionally, the increased gene levels of LDHA, CD147, MCT4, PI3K, AKT, mTOR and HIF-1α in model group were down-regulated by WPX decoction, while miRNA-34a expression was decreased and up-regulated by WPX decoction. The significantly increased protein levels of LDHA, CD147, MCT4, PI3K, AKT, mTOR and HIF-1α induced by MNNG were attenuated in rats treated with WPX decoction. In brief, the findings of this study imply that abnormal glycolysis in MNNG-induced GPL rats was relieved by WPX decoction via regulation of the expressions of LDHA, CD147, HIF-1α, MCT4, PI3K, AKT, mTOR and miRNA-34a.

Weipixiao attenuate early angiogenesis in rats with gastric precancerous lesions.

BACKGROUND: Angiogenesis is a pathobiological hallmark of gastric cancer. However, rare studies focus on angiogenesis in gastric precancerous lesions (GPL). Weipixiao (WPX), a Chinese herbal preparation, is proved clinically effective in treating GPL. Here, we evaluated WPX's anti-angiogenic potential for GPL, and also investigated the possibility of its anti-angiogenic mechanisms. METHODS: HPLC analysis was applied to screen the major chemical components of WPX. After modeling N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-induced GPL in male Sprague-Dawley rats, different doses of WPX were administrated orally for 10 weeks. Next, we performed histopathological examination using routine H&E staining and HID-AB-PAS staining. In parallel, we assessed angiogenesis revealed by microvessel density (MVD) using CD34 immunostaining, and subsequently observe microvessel ultrastructure in gastric mucosa under Transmission Electron Microscope. Finally, we detect expression of angiogenesis-associated markers VEGF and HIF-1α using immunohistochemistry. Moreover, mRNA expressions of ERK1, ERK2, Cylin D1 as well as HIF-1α in gastric mucosa were determined by quantitative real-time reverse transcription- polymerase chain reaction. RESULTS: We observed the appearance of active angiogenesis in GPL rats, and demonstrated that WPX could reduce microvascular abnormalities and attenuate early angiogenesis in most of GPL specimens with a concomitant regression of most intestinal metaplasia (IM) and a portion of gastric epithelial dysplasia (GED). In parallel, WPX could suppress HIF-1α mRNA expression (P < 0.01) as well as protein expression (although without statistical significance), and could markedly inhibit VEGF protein expression in GPL rats. Mechanistically, WPX intervention, especially at low dose, caused a significant decrease in the ERK1 and Cylin D1 mRNA levels. However, WPX might probably have no regulatory effect on ERK2 amplification. CONCLUSIONS: WPX could attenuate early angiogenesis and temper microvascular abnormalities in GPL rats. This might be partly achieved by inhibiting on the angiogenesis-associated markers HIF-1α and VEGF, and on the ERK1/Cylin D1 aberrant activation.

Astragaloside IV reverses MNNG-induced precancerous lesions of gastric carcinoma in rats: Regulation on glycolysis through miRNA-34a/LDHA pathway.

This study was designed to investigate the precancerous lesions of gastric carcinoma (PLGC)-reversing mechanisms of astragaloside IV (ASIV) in N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-induced PLGC rats. All rats were sacrificed after 10-week treatment. Gastric tissue was analyzed by using histopathology and electron microscope. To be fully evidenced, LDHA, p53, TIGAR, MCT1, MCT4, HIF-1α, CD147, and miRNA-34a were detected by Western blotting and Real-time Quantitative polymerase chain reaction (RT-qPCR). As histopathology and electron microscope showed, it can be clearly observed that the area of dysplasia was reduced in ASIV groups, indicating that MNNG-induced PLGC was markedly reversed by ASIV. Moreover, compared with model group, a significant decrease in gene expressions of LDHA, MCT1, MCT4, HIF-1α, CD147, and TIGAR was observed whereas miRNA-34a level was increased in ASIV groups. A significant up-regulation induced by MNNG in protein levels of LDHA, MCT1, MCT4, HIF-1α, and CD147 was attenuated in rats treated with ASIV. In contrast, the decreased expression of TIGAR was restored by ASIV. Interestingly, up-regulation of p53 expression induced by MNNG was further increased in ASIV groups. In brief, these results implied that abnormal glycolysis was relieved by ASIV via regulation of the expressions of LDHA, p53, TIGAR, MCT1, MCT4, HIF-1α, CD147, and miRNA-34a.

Weipixiao ameliorates gastric precancerous lesions in a rat's model by regulating GSK3¦Â and C-myc.

OBJECTIVE: To investigate the mechanism underlying the action of Weipixiao (WPX) in a rat's model with ameliorating gastric precancerous lesions (GPL). METHODS: HPLC analysis was performed to identify the chemical constituents of WPX preparation. Sprague- Dawley rats were randomly assigned into control group, model group, vitacoenzyme group, high-dose WPX group (H-WPX), medium-dose WPX group (M-WPX) and low-dose WPX group (L-WPX). After modeling, the treated rats were administrated WPX or vitacoenzyme intragastrically for consecutive 10 weeks. Gene and protein expressions of GSK3¦Â, C-myc, Cylin E were evaluated by quantitative real-time reverse transcription-polymerase chain reaction (RT-qPCR) and immunohistochemistry, respectively. RESULTS: WPX could efficiently attenuate the pathological alterations of ""non-progressive GPL"" in rats. As expected, mRNA and protein levels of C-myc and Cylin E were up-regulated in model rats, while GSK3¦Â expression down-regulated (P < 0.01). WPX treatment, especially at low dose, could significantly down-regulate the mRNA as well as protein levels of C-myc, and could lead to remarkable up-regulation of mRNA and protein levels of GSK3¦Â in GPL rats (P < 0.05). However, no significant changes were observed in WPX-treated rats. CONCLUSION: Our findings suggested that WPX-mediated attenuation of GPL pathological alterations might be due to its regulatory effect on the expressions of GSK3¦Â and C-myc, and on the dysregulation of Wnt/GSK3¦Â pathway.

Preparation and deproteinization of garlic polysaccharide.

Preparation and deproteinization methods of polysaccharide from garlic (Allium sativum L.) were studied. The crude polysaccharide was prepared by the method of hot water extraction. The percentages of deproteinization and polysaccharide loss were compared as indexes using the hydrochloric acid (HCl) method, trichloroacetic acid method, NaCl method and CaCl(2) method, respectively. The infrared spectra analysis and content analysis showed that the HCl method exhibited the highest percentage of deproteinization, and a little higher percentage of polysaccharide loss than the other three methods. The CaCl(2) method exceeded NaCl method in deproteinization.

Synthesis, biological evaluation of chrysin derivatives as potential immunosuppressive agents.

A series of novel chrysin derivatives was firstly synthesized and evaluated on their immunosuppressive activity in the search for potential immunosuppressive agents. Among them, compounds 5c displayed the most potent immunosuppressive inhibitory activity with IC(50) of 0.78 µM, which was comparable to that of cyclosporin A (IC(50) = 0.06 µM). The preliminary mechanism of compound 5c inhibition effects was also detected by flow cytometry (FCM), and the compound exerted immunosuppressive activity via inducing the apoptosis of activated lymph node cells in a dose dependent manner. Furthermore, the estimated LD(50) (in mg/kg) in vivo of compound 5c is 738.2, which indicated that compound 5c was low toxic.

Si-Ni-San, a traditional Chinese prescription, and its active ingredient glycyrrhizin ameliorate experimental colitis through regulating cytokine balance.

Si-Ni-San, a traditional Chinese medicinal formula, exerts an important function in the treatment of inflammatory bowel diseases based upon thousands of years of clinical practice, but the underlying mechanism is still unclear. In this study, we investigated the therapeutic potential of Si-Ni-San and its ingredient glycyrrhizin in trinitrobenzene sulfonic acid (TNBS)-induced experimental colitis in mice, a well-characterized murine model for Crohn's disease. Si-Ni-San and glycyrrhizin significantly ameliorated TNBS-induced colitis with reduced mortality and recovery of body weights. In addition, Si-Ni-San and glycyrrhizin dose-dependently decreased macroscopic inflammation scores, microscopic histological scores, and myeloperoxidase activity. Furthermore, Si-Ni-San and glycyrrhizin caused a decrease in pro-inflammatory cytokines including IFN-gamma, IL-12, TNF-alpha and IL-17 and an increase in regulatory cytokine IL-10 in colon of the mice. It should be noticed the therapeutic effect of Si-Ni-San at 450 mg/kg was much better than that of its contained content of glycyrrhizin at 10 mg/kg. In conclusion, Si-Ni-San and glycyrrhizin significantly ameliorated TNBS-induced colitis in mice through regulating pro- and anti-inflammatory cytokine production.

Dissection of the role of paeoniflorin in the traditional Chinese medicinal formula Si-Ni-San against contact dermatitis in mice.

AIMS: The roles of specific active ingredients in Chinese medicinal formulas have not been clearly elucidated. In this study, we selectively deleted and replenished paeoniflorin from Si-Ni-San, a traditional Chinese prescription, and aimed to identify the molecular basis of how paeoniflorin exerted its effect in Si-Ni-San. MAIN METHODS: Contact dermatitis was induced in mice with picryl chloride. Paeoniflorin was selectively deleted from Si-Ni-San by an immunoaffinity column. Quantitative real-time PCR, western blot, and enzyme-linked immunosorbent assay were used in this study. KEY FINDINGS: Both Si-Ni-San and paeoniflorin significantly reduced ear swelling in mice while the paeoniflorin-deleted Si-Ni-San (Si-Ni-San(PF-)) showed little ameliorative effect. In lipopolysaccharide-evoked macrophages, Si-Ni-San and paeoniflorin markedly inhibited tumor necrosis factor-alpha production, cyclooxygenase-2 activity, as well as extracellular signal-regulated kinase 1/2 phosphorylation while Si-Ni-San(PF-) exhibited no or slight inhibitory effect. Furthermore, the inhibitory effect on the production of tumor necrosis factor-alpha reappeared when different proportions of paeoniflorin were replenished in Si-Ni-San(PF-). In addition, the expression of macrophage migration inhibitory factor in T cells, rather than macrophages, was significantly inhibited by Si-Ni-San, but not Si-Ni-San(PF-). Our data indicate paeoniflorin is the principal component of Si-Ni-San, exerting negative regulation on the function of macrophages in contact dermatitis. SIGNIFICANCE: The present study suggests that dissecting the role of specific constituents in medicinal formulas through selective deletion and replenishment may be a useful strategy in recognizing and validating an active ingredient in traditional Chinese medicine.